Determination of intestinal microbioma in patients with diarrhea associated with Clostridium difficile infection acquired in Intensive Care Unit and community.

Clostridium difficile (CD) has been identified as the leading cause of diarrhea associated with overuse of antibiotics. CD infection (ICD) ranges from mild to severe diarrhea to pseudomembranous colitis and toxic megacolon, and can even be lethal. The ICD has increased its incidence, severity and mortality, due to the emergence of hypervirulent strains and the acquisition of antibiotic resistance, which added to the high frequency of recurrences, has made it a priority at the level of health research. Populations exposed to antibiotics are the most affected by the ICD, particularly patients in Intensive Care Units (ICU), where this pathogen is one of the five most frequent infections.

The main factor associated with the success of the ICD is the deregulation of the host's intestinal microbiota, involved in the modulation of metabolic and defense (immune and barrier) functions. This deregulation is generally derived from the use of antibiotics, an event required for the germination of CD spores, which leads to the replication of vegetative forms producing clostridial toxins (the main virulence factor of this pathogen). Although the bacterial component is determinant for the adequate function of the microbiota, recently other groups of organisms, such as viruses and eukaryotes, including protists and helminths, have been described as fundamental members of microbial communities.

In addition to the composition of this microbiota, it has been widely reported that the presence of mobile genetic elements present at the intestinal level is related to the mobilization of clinically important loci, especially genes associated with antibiotic resistance.

For these reasons, this study is aimed at determining the communities that compose the microbiota (bacterioma, fungoma and eukaryoma) of patients with diarrhea associated to the ICD acquired in the ICU and in the community, comparing the distribution patterns of these species and their population structure at the genetic level. Additionally, it is sought to evaluate the presence of circulating mobile genetic elements in these groups of patients, possibly involved in the alteration of the dynamics of microbial communities.

The project involves the processing of stool samples from patients with diarrhea at intra-hospital and community level, collected during the execution of a previous project. These samples were subjected to identification of the ICD as criteria for the selection of 100 samples, which included: 60 positive samples for the ICD (30 of them from ICU and another 30 from the community) and 40 negative samples for the ICD (), 20 from each population studied.

DNA and RNA will be extracted and subjected to high performance sequencing using the Illumina Hi Seq platform. From these data, the taxonomic identification of the species will be carried out and the abundance, diversity and population structure of the identified taxonomic units will be evaluated. In addition, the presence of mobile genetic elements associated with antibiotic resistance will be evaluated.

This study is a pioneer in the implementation of high-performance sequencing techniques for the description of the intestinal microbiota associated with the ICD acquired at the intra-hospital and community level. The results of this study seek to provide information that in the future favors the improvement of prevention strategies, timely diagnosis and appropriate treatment, which can reduce complications and death of patients from this infection.